The present invention relates to inhibitors of cyclin dependent kinase and their use in the treatment of proliferative diseases.
Normal cellular proliferation is stringently regulated by a series of proteins that constitute the cell cycle machinery. Proteins that play a key role in controlling cell cycle progression are the cyclin dependent kinases (CDKs). CDKs are serine/threonine protein kinases that are the driving force behind the cell cycle and cell proliferation. The active CDK enzyme is a multi-subunit complex composed of at least one catalytic (CDK) subunit and one regulatory (cyclin) subunit. See, Brooks and La Thangue, DDT, 4, 455-464 (1999). It has been found that inhibitors of CDK activity are effective for the treatment of proliferative diseases (e.g. cancer). See, Webster and Kimball, Emerging Drugs, 5, 45-59 (2000).
The present invention is directed to the use of compounds of formula 
and pharmaceutically acceptable salts thereof as inhibitors of cyclin dependent kinases. As used in formula I, and throughout the specification, the symbols have the following meanings:
R1 is hydrogen, aryl or lower alkyl;
R2 and R4 are each independently alkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, cycloalkyl, cycloalkylalkyl, heterocycloalkyl or heterocycloalkylalkyl;
R3 is hydrogen or lower alkyl; and
n is an integer of 0, 1 or 2.
The compounds of formula I are protein kinase inhibitors and are useful in the treatment and prevention of proliferative diseases, for example, cancer, inflammation and arthritis. They may also be useful in the treatment of neurodegenerative diseases such Alzheimer""s disease, cardiovascular diseases, viral diseases and fungal diseases.
The present invention provides for a method of using compounds of formula I as inhibitors of cyclin dependent kinases, which are active in the treatment of proliferative diseases, such as for example, but not limited to, cancer, Alzheimer""s disease, arthritis, inflammation, and cardiovascular disease. The present invention also contemplates pharmaceutical compositions employing such compounds.
Listed below are definitions of various terms used to describe the compounds of the instant invention. These definitions apply to the terms as they are used throughout the specification (unless they are otherwise limited in specific instances) either individually or as part of a larger group.
It should be noted that any heteroatom with unsatisfied valences is assumed to have hydrogen atoms sufficient to satisfy the valences.
Carboxylate anion refers to a negatively charged group xe2x80x94COOxe2x80x94.
The term xe2x80x9calkylxe2x80x9d or xe2x80x9calkxe2x80x9d refers to a monovalent alkane (hydrocarbon) derived radical containing from 1 to 12 carbon atoms unless otherwise defined. The term xe2x80x9clower alkylxe2x80x9d refers to an alkyl group of 1 to 6 carbon atoms. An alkyl group is an optionally substituted straight, branched or cyclic saturated hydrocarbon group. When substituted, alkyl groups may be substituted with up to four substituent groups, as defined below, at any available point of attachment. When the alkyl group is said to be substituted with an alkyl group, this is used interchangeably with xe2x80x9cbranched alkyl groupxe2x80x9d. Exemplary unsubstituted such groups include methyl, ethyl, propyl, isopropyl, n-butyl, t-butyl, isobutyl, pentyl, hexyl, isohexyl, heptyl, 4,4-dimethylpentyl, octyl, 2,2,4-trimethylpentyl, nonyl, decyl, undecyl, dodecyl, and the like. Exemplary substituent groups (or substituents) may include, but are not limited to, one or more of the following groups: halo (such as F, Cl, Br or I), haloalkyl (such as CCl3 or CF3), alkoxy, aryloxy, alkyl S(O)m (m=0, 1, 2), aryl S(O)m (m=0, 1, 2), hydroxy, carboxy (xe2x80x94COOH), alkyloxycarbonyl (xe2x80x94COORxe2x80x2), alkylcarbonyloxy (xe2x80x94OCORxe2x80x2), amino (xe2x80x94NH2), quaternary nitrogen, carbamoyl (xe2x80x94NHCOORxe2x80x2xe2x80x94 or xe2x80x94OCONHRxe2x80x2xe2x80x94), urea (xe2x80x94NHCONHRxe2x80x2xe2x80x94), thiol (xe2x80x94SH), cyano or nitro. Alkyl groups as defined may also comprise one or more carbon to carbon double bonds or one or more carbon to carbon triple bonds.
The term xe2x80x9calkenylxe2x80x9d refers to a hydrocarbon radical straight, branched or cyclic containing from 2 to 12 carbon atoms and at least one carbon to carbon double bond.
The term xe2x80x9calkynylxe2x80x9d refers to a hydrocarbon radical straight, branched or cyclic containing from 2 to 12 carbon atoms and at least one carbon to carbon triple bond.
Cycloalkyl is a type of alkyl containing from 3 to 15 carbon atoms, without alternating or resonating double bonds between carbon atoms. It may contain from 1 to 4 rings. Exemplary unsubstituted such groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, etc. Exemplary substituents include one or more of the following groups: halogen, alkyl, alkoxy, alkyl, hydroxy, amino, nitro, cyano, thiol and/or alkylthio.
The terms xe2x80x9calkoxyxe2x80x9d or xe2x80x9calkylthioxe2x80x9d, as used herein, denote an alkyl group as described above bonded through an oxygen linkage (xe2x80x94Oxe2x80x94) or a sulfur linkage (xe2x80x94Sxe2x80x94), respectively.
The term xe2x80x9calkyloxycarbonylxe2x80x9d, as used herein, denotes an alkoxy group bonded through a carbonyl group. An alkoxycarbonyl radical is represented by the formula xe2x80x94C(O)OR, where the R group is a straight or branched C1-6 alkyl group.
The term xe2x80x9calkylcarbonylxe2x80x9d refers to an alkyl group bonded through a carbonyl group.
The term xe2x80x9calkylcarbonyloxyxe2x80x9d, as used herein, denotes an alkylcarbonyl group which is bonded through an oxygen linkage.
The term xe2x80x9carylalkylxe2x80x9d, as used herein, denotes an aromatic ring bonded through an alkyl group as described above.
The term xe2x80x9carylxe2x80x9d refers to monocyclic or bicyclic aromatic rings, e.g., phenyl, substituted phenyl and the like, as well as groups which are fused, e.g., napthyl, phenanthrenyl and the like. An aryl group thus contains at least one ring having at least 6 atoms, with up to five such rings being present, containing up to 22 atoms therein, with alternating (resonating) double bonds between adjacent carbon atoms or suitable heteroatoms. Aryl groups may optionally be substituted with one or more groups including, but not limited to, halogen, alkyl, alkoxy, hydroxy, carboxy, carbamoyl, alkyloxycarbonyl, nitro, trifluoromethyl, amino, cycloalkyl, cyano, alkylS(O)m (m=0, 1, 2), or thiol.
The term xe2x80x9cheteroarylxe2x80x9d refers to a monocyclic aromatic hydrocarbon group having 5 or 6 ring atoms, or a bicyclic aromatic group having 8 to 10 atoms, containing at least one heteroatom, O, S, or N, in which a carbon or nitrogen atom is the point of attachment, and in which one or two additional carbon atoms is optionally replaced by a heteroatom selected from O or S, and in which from 1 to 3 additional carbon atoms are optionally replaced by nitrogen heteroatoms, said heteroaryl group being optionally substituted as described herein. Additional nitrogen atoms may be present together with the first nitrogen and oxygen or sulfur. Exemplary heteroaryl groups include the following: thienyl, furyl, pyrrolyl, pyridinyl, imidazolyl, pyrrolidinyl, piperidinyl, thiazolyl, oxazolyl, triazolyl, pyrazolyl, isoxazolyl, isothiazolyl, pyrazinyl, pyridazinyl, pyrimidinal, triazinylazepinyl, indolyl, isoindolyl, quinolinyl, isoquinolinyl, benzothiazolyl, benzoxazolyl, benzimidazolyl, benzoxadiazolyl, benzofurazanyl and tetrahydropyranyl. Exemplary substituents include one or more of the following: halo, alkyl, alkoxy, hydroxy, cycloalkyl, nitro, cyano, amino, alkylS(O)m (m=0, 1, 2), or thiol. The term xe2x80x9cheteroarylalkylxe2x80x9d, as used herein denotes a heteroaryl ring bonded through an alkyl group as described hereinabove.
The term xe2x80x9cheteroaryliumxe2x80x9d refers to heteroaryl groups bearing a quaternary nitrogen atom and thus a positive charge.
The term xe2x80x9cheterocycloalkylxe2x80x9d refers to a cycloalkyl group (nonaromatic) in which one of the carbon atoms in the ring is replaced by a heteroatom selected from O, S or N, and in which up to three additional carbon atoms may be replaced by said heteroatoms. In addition, the sulfur may be oxidated to the sulfone (xe2x80x94SO2xe2x80x94) or sulfoxide (xe2x80x94SOxe2x80x94) and the nitrogen may be quaternary.
The term xe2x80x9cquaternary nitrogenxe2x80x9d refers to a tetravalent positively charged nitrogen atom including, e.g., the positively charged nitrogen in a tetraalkylammonium group (e.g., tetramethylammonium, N-methylpyridinium), the positively charged nitrogen in protonated ammonium species (e.g., trimethylhydroammonium, N-hydropyridinium), the positively charged nitrogen in amine N-oxides (e.g. N-methyl-morpholine-N-oxide, pyridine-N-oxide), and the positively charged nitrogen in an N-amino-ammonium group (e.g., N-aminopyridinium).
The term xe2x80x9cheteroatomxe2x80x9d means O, S, P or N, selected on an independent basis.
The term xe2x80x9chalogenxe2x80x9d or xe2x80x9chaloxe2x80x9d refers to chlorine, bromine, fluorine or iodine.
The term xe2x80x9cPMBxe2x80x9d refers to para-methoxybenzyl.
When a functional group is termed xe2x80x9cprotectedxe2x80x9d, this means that the group is in modified form to preclude undesired side reactions at the protected site. Suitable protecting groups for the compounds of the present invention will be recognized from the present application taking into account the level of skill in the art, and with reference to standard textbooks, such as Greene, T. W. et al., Protective Groups in Organic Synthesis, Wiley, N.Y. (1991).
Suitable examples of salts of the compounds according to the invention with inorganic or organic acids are hydrochloride, hydrobromide, sulfate, phosphate. Salts which are unsuitable for pharmaceutical uses but which can be employed, for example, for the isolation or purification of free compounds I or their pharmaceutically acceptable salts, are also included.
All stereoisomers of the compounds of the instant invention are contemplated, either in admixture or in pure or substantially pure form. The definition of the compounds according to the invention embraces all possible stereoisomers and their mixtures. It very particularly embraces the racemic forms and the isolated optical isomers having the specified activity. The racemic forms can be resolved by physical methods, such as, for example, fractional crystallization, separation or crystallization of diastereomeric derivatives or separation by chiral column chromatography. The individual optical isomers can be obtained from the racemates by conventional methods, such as, for example, salt formation with an optically active acid followed by crystallization.
It should be understood that the present invention includes prodrug forms of the compounds of formula I. Various forms of prodrugs are well known in the art. Examples of such prodrug derivatives are provided in the following references:
(a) Design of Prodrugs, edited by H. Bundgaard (Elsevier, 1985); and Methods in Enzymology, Vol. 42, pp. 309-396, edited by K. Widder et al., (Academic Press, 1985);
(b) A Textbook of Drug Design and Development, edited by Krosgaard-Larsen and H. Bundgaard, Chapter 5, xe2x80x9cDesign and Application of Prodrugs,xe2x80x9d by H. Bundgaard, pp. 113-191 (1991);
(c) H. Bundgaard, Advanced Drug Delivery Reviews, 8, pp. 1-38 (1992);
(d) H. Bundgaard et al., Journal of Pharmaceutical Sciences, 77, 285 (1988); and
(e) N. Kayeka et al., Chem. Phar. Bull., 32, 692 (1984).
It should be understood that solvates (e.g., hydrates) of the compounds of formula I are also within the scope of the present invention. Methods of solvation are generally known in the art. Accordingly, the compounds of the instant invention may be in the free or hydrate form, and may be obtained by methods exemplified by the following schemes.
Compounds of formula I for use as inhibitors of cyclin dependent kinases can be prepared by reacting a compound of formula II 
with a base such as sodium hydroxide or sodium carbonate to give a free base of formula IIa 
Compound IIa is reacted with a compound of formula III 
at an elevated temperature, preferably about 130xc2x0 C., under reduced pressure to obtain compounds of formula IV 
The starting compound of formula II, where R1 is hydrogen is prepared by reacting acrylonitrile with hydrazine hydrate in a solvent such as THF, followed by addition of p-methoxybenzaldehyde. The compound which results from this reaction is then reacted with a mixture of sodium n-butoxide in n-butanol followed by HCl to provide the key early intermediate of compound II.
Compounds of formula IV are then reacted at an elevated temperature, preferably from about 220xc2x0 C. to 260xc2x0 C., in the presence of a solvent such as diphenyl ether to obtain a compound of formula V 
Compound V is then reacted with an aqueous base such as aqueous sodium hydroxide or aqueous potassium hydroxide at an elevated temperature, preferably about 95xc2x0 C., to obtain a compound of formula VI 
Compound VI is stirred at an elevated temperature, preferably about 230xc2x0 C., to obtain a compound of formula VII 
The formula VII compound is reacted with bromine in the presence of a solvent such as ethanol, preferably at a temperature of about 0xc2x0 C., to obtain a compound of formula VIII 
Compound VIII is then reacted with phosphorous oxychloride at a temperature from about 25xc2x0 to 130 xc2x0 C. to obtain a compound of formula IX 
Compound IX is reacted with a compound of formula X
MOR4 xe2x80x83xe2x80x83(X) 
wherein M is an alkali metal such as sodium or potassium, and R4 is as described hereinabove in the presence of a solvent such as tetrahydrofuran, R4OH and the like at a temperature from about 25xc2x0 C. to 90xc2x0 C. to form a compound of formula XI 
The formula XI compound is reacted with an organolithium reagent such as n-butyllithium in a solvent such as tetrahydrofuran, ether and the like at a temperature from about xe2x88x9290xc2x0 to xe2x88x9220xc2x0 C. followed by treatment with a compound of formula XII
R2SSR2 xe2x80x83xe2x80x83(XII) 
wherein R2 is as described hereinabove to obtain a compound of formula XIII 
Compounds of formula XIII are then reacted with:
(1) an acid, such as trifluoroacetic acid (TFA), at a temperature from about 25xc2x0 C. to 90xc2x0 C., or
(2) hydrogen in the presence of a catalyst, such as palladium on carbon, to obtain compounds of formula I wherein n is 0 
Compounds of formula I wherein n is 1 or 2 may be prepared by oxidizing compounds of formula I wherein n is 0 with an oxidizing agent such as m-chloroperoxybenzoic acid (mCPBA) and the like in the presence of a solvent such as dichloromethane. Generally one equivalent of the oxidizing agent is used to obtain compounds of formula I wherein n is 1, and two or more equivalents of the oxidizing agent are used to obtain compounds of formula I wherein n is 2. 
Intermediates of this invention may also be prepared by processes disclosed in U.S. Pat. Nos. 3,828,057, 3,966,746, 3,979,399, and 3,985,757 which are incorporated by reference herein. More specifically, intermediates of formula II may be prepared by procedures described in Hoehn, H., Z. Chem. 10, pp. 386-388 (1970).
Compounds of formulas II and III are commercially available or may be prepared by methods known to one of ordinary skill in the art.
All other compounds may be prepared by modification of the procedures described herein.
The preferred compounds of formula I are those wherein:
R1 and R3 are hydrogen;
R2 is aryl or heteroaryl;
R4 is alkyl, cycloalkyl or cycloalkylalkyl; and
n is an integer of 0, 1 or 2.
More preferred compounds of formula I are those wherein:
R1 and R3 are hydrogen;
R2 is phenyl, 4-bromo-2,6-difluorophenyl, 4-chloro-2,6-difluorophenyl, 2,4,6-trifluorophenyl or 4-methyl-2,6-difluorophenyl;
R4 is lower alkyl; and
n is 1.
The compounds according to the invention have pharmacological properties; in particular, the compounds of formula I are inhibitors of protein kinases such as the cyclin dependent kinases (cdks), for example, cdc2 (cdk1), cdk2, and cdk4. The novel compounds of formula I are expected to be useful in the therapy of proliferative diseases such as cancer, inflammation, arthritis, Alzheimer""s disease and cardiovascular disease. These compounds may also be useful in the treatment of topical and systemic fungal infections.
More specifically, the compounds of formula I are useful in the treatment of a variety of cancers, including (but not limited to) the following:
carcinoma, including that of the bladder, breast, colon, kidney, liver, lung, ovary, pancreas, stomach, cervix, thyroid, prostate, and skin;
hematopoietic tumors of lymphoid lineage, including acute lymphocytic leukemia, B-cell lymphoma, and Burkett""s lymphoma;
hematopoietic tumors of myeloid lineage, including acute and chronic myelogenous leukemias and promyelocytic leukemia;
tumors of mesenchymal origin, including fibrosarcoma and rhabdomyosarcoma; and
other tumors, including melanoma, seminoma, teratocarcinoma, osteosarcoma, neuroblastoma and glioma.
Due to the key role of cdks in the regulation of cellular proliferation in general, inhibitors could act as reversible cytostatic agents which may be useful in the treatment of any disease process which features abnormal cellular proliferation, e.g., neuro-fibromatosis, atherosclerosis, pulmonary fibrosis, arthritis, psoriasis, glomerulonephritis, restenosis following angioplasty or vascular surgery, hypertrophic scar formation, inflammatory bowel disease, transplantation rejection, angiogenesis, and endotoxic shock.
Compounds of formula I may also be useful in the treatment of Alzheimer""s disease, as suggested by the recent finding that cdk5 is involved in the phosphorylation of tau protein (J. Biochem, 117, 741-749 (1995)).
Compounds of formula I may also act as inhibitors of other protein kinases, e.g., protein kinase C, her2, rafl, MEK1, MAP kinase, EGF receptor, PDGF receptor, IGF receptor, PI3 kinase, wee1 kinase, Src, Ab1, VEGF, and 1ck, and thus be effective in the treatment of diseases associated with other protein kinases.
Compounds of formula I also induce or inhibit apoptosis, a physiological cell death process critical for normal development and homeostasis. Alterations of apoptotic pathways contribute to the pathogenesis of a variety of human diseases. Compounds of formula I, as modulators of apoptosis, will be useful in the treatment of a variety of human diseases with abberations in apoptosis including cancer (particularly, but not limited to, follicular lymphomas, carcinomas with p53 mutations, hormone dependent tumors of the breast, prostate and ovary, and precancerous lesions such as familial adenomatous polyposis), viral infections (including, but not limited to, herpesvirus, poxvirus, Epstein-Barr virus, Sindbis virus and adenovirus), autoimmune diseases (including, but not limited to, systemic lupus, erythematosus, immune mediated glomerulonephritis, rheumatoid arthritis, psoriasis, inflammatory bowel diseases, and autoimmune diabetes mellitus), neurodegenerative disorders (including, but not limited to, Alzheimer""s disease, AIDS-related dementia, Parkinson""s disease, amyotrophic lateral sclerosis, retinitis pigmentosa, spinal muscular atrophy and cerebellar degeneration), AIDS, myelodysplastic syndromes, aplastic anemia, ischemic injury associated myocardial infarctions, stroke and reperfusion injury, arrhythmia, atherosclerosis, toxin-induced or alcohol induced liver diseases, hematological diseases (including, but not limited to, chronic anemia and aplastic anemia), degenerative diseases of the musculoskeletal system (including, but not limited to, osteoporosis and arthritis), aspirin-sensitive rhinosinusitis, cystic fibrosis, multiple sclerosis, kidney diseases, and cancer pain.
The compounds of this invention may also be useful in combination with known anti-cancer treatments such as radiation therapy or with cytostatic and cytotoxic agents, such as, for example, but not limited to, DNA interactive agents, such as cisplatin or doxorubicin; inhibitors of farnesyl protein transferase, such as those described in U.S. Pat. No. 6,011,029, topoisomerase II inhibitors, such as etoposide; topoisomerase I inhibitors, such as CPT-11 or topotecan; tubulin stabilizing agents, such as paclitaxel, docetaxel or the epothilones; hormonal agents, such as tamoxifen; thymidilate synthase inhibitors, such as 5-fluorouracil; and antimetabolites, such as methoxtrexate; antiangiogenic agents, such as angiostatin or endostatin; and kinase inhibitors, such as her2 specific antibodies. The formula I compounds of this invention may also be useful in combination with modulators of p53 transactivation. In addition, the formula I compounds may be used for treating chemotherapy-induced alopecia, chemotherapy-induced thrombocytopenia, chemotherapy-induced leukopenia or mucocitis. In the treatment of chemotherapy-induced alopecia, the formula I compound is preferably topically applied in the form of a medicament such as a gel, shampoo, aerosol, dust, cream, ointment, solution, dispersion or paste.
If formulated as a fixed dose, such combination products employ the compounds of this invention within the dosage range described below and the other pharmaceutically active agent within its approved dosage range. For example, the cdc2 inhibitor olomucine has been found to act synergistically with known cytotoxic agents in inducing apoptosis (J. Cell Sci., 108, 2897 (1995)). Compounds of formula I may be used sequentially with known anti-cancer or cytotoxic agents when a combination formulation is inappropriate.
cdc2/cyclin B1 kinase activity was determined by monitoring the incorporation of 32p into histone HI. The reaction consisted of 50 ng baculovirus expressed GST-cdc2, 75 ng baculovirus expressed GST-cyclin B 1, 1 xcexcg histone HI (Boehringer Mannheim), 0.2 xcexcCi of 32p xcex3-ATP and 25 xcexcM ATP in kinase buffer (50 mM Tris, pH 8.0, 10 mM MgCl2, 1 mM EGTA, 0.5 mM DTT). The reaction was incubated at 30 xc2x0 C. for 30 minutes and then stopped by the addition of cold trichloroacetic acid (TCA) to a final concentration of 15% and incubated on ice for 20 minutes. The reaction was harvested onto GF/C unifilter plates (Packard) using a Packard Filtermate Universal harvester, and the filters were counted on a Packard TopCount 96-well liquid scintillation counter (Marshak, D. R., Vanderberg, M. T., Bae, Y. S., Yu, I. J., J. Cellular Biochemistry, 45, 391-400 (1991), incorporated by reference herein).
cdk2/cyclin E kinase activity was determined by monitoring the incorporation of 32p into the retinoblastoma protein. The reaction consisted of 2.5 ng baculovirus expressed GST-cdk2/cyclin E, 500 ng bacterially produced GST-retinoblastoma protein (aa 776-928), 0.2 xcexcCi 32P xcex3-ATP and 25 xcexcM ATP in kinase buffer (50 mM Hepes, pH 8.0, 10 mM MgCl2, 5 mM EGTA, 2 mM DTT). The reaction was incubated at 30 xc2x0 C. for 30 minutes and then stopped by the addition of cold trichloroacetic acid (TCA) to a final concentration of 15% and incubated on ice for 20 minutes. The reaction was harvested onto GF/C unifilter plates (Packard) using a Packard Filtermate Universal harvester, and the filters were counted on a Packard TopCount 96-well liquid scintillation counter.
cdk4/cyclin D1 kinase activity was determined by monitoring the incorporation of 32p in to the retinoblastoma protein. The reaction consisted of 165 ng baculovirus expressed as GST-cdk4, 282 ng bacterially expressed as S-tag cyclin D1, 500 ng bacterially produced GST-retinoblastoma protein (aa 776-928), 0.2 xcexcCi 32P xcex3-ATP and 25 xcexcM ATP in kinase buffer (50 mM Hepes, pH 8.0, 10 mM MgCl2, 5 mM EGTA, 2 mM DTT). The reaction was incubated at 30xc2x0 C. for 1 hour and then stopped by the addition of cold trichloroacetic acid (TCA) to a final concentration of 15% and incubated on ice for 20 minutes. The reaction was harvested onto GF/C unifilter plates (Packard) using a Packard Filtermate Universal harvester, and the filters were counted on a Packard TopCount 96-well liquid scintillation counter (Coleman, K. G., Wautlet, B. S., Morissey, D, Mulheron, J. G., Sedman, S., Brinkley, P., Price, S., Webster, K. R. (1997) Identification of CDK4 Sequences involved in cyclin D, and p16 binding. J. Biol. Chem. 272,30:18869-18874, incorporated by reference herein).
Further subject matter of the invention also includes pharmaceuticals for use as described above including controlling cancer, inflammation and arthritis, which contain at least one compound of formula I as defined above or at least one of its pharmacologically acceptable acid addition salts, and the use of a compound of formula I as defined above for the preparation of a pharmaceutical having activity against proliferative diseases as described previously including against cancer, inflammation and/or arthritis.
The following examples are illustrative rather than limiting. It should be understood that there may be other embodiments which fall within the spirit and scope of the invention as defined by the claims appended hereto.